The proposed research is designed to explore several aspects of presynaptic nerve terminal function; pinched-off terminals (synaptosomes) will be employed in these studies. One series of experiments focuses on the mechanism underlying "active" calcium extrusion across the nerve terminal plasma membrane. Previous work indicates that an exchange of sodium for calcium may be involved, and that energy from the sodium electrochemical gradient may provide energy for this Na-Ca countertransport. The kinetic details of this exchange will be examined in order to determine whether the "carrier" mediates sequential or simultaneous ion counter-transport. A determined effort will be made to 1) isolate the "carrier" from synaptic plasma membranes and 2) reconstitute the "carrier" into lipid bilayer vesicles. A second series of experiments focuses on calcium sequestration within the nerve terminals. Thie sequestration is ATP-dependent, but apparenly does not involve mitochondria because it is unaffected by typical mitochondrial poisons. We will attempt to determine 1) whether or not ATP is hydrolyzed when Ca is sequestered, and 2) which organelles are the locus of this Ca storage. The Ca-sensitive metallochrome, arsenazo III, will be employed in order to evaluate the kinetics of Ca sequestration and to determine the concentration at which Ca is buffered inside the terminals. Synaptic vesicle exocytosis-retrieval-recycling processes are the focus of a third set of experiments. The main aims here are 1) to obtain direct evidence for synaptic vesicle-plasma membrane fusion, and 2) to compare the protein composition of the normal synaptic vesicles (in resting terminals) and the retrieved vesicles - to determine whether or not plasma membrane components (e.g., Na plus K-dependent ATPase) are present in the membranes of the newly-retrieved vesicles. The fourth series of experiments is designed to test the influence of the membrane potential on the carrier-mediated (Na-dependent) influx and efflux of gamma-aminobutyric acid (GABA). The data will be used to test the applicability of the currently-popular absolute-reaction-rate-theory model (the "barrier model" of Hall et al., J. Membrane Biol., 11: 75, 1973) to this transport process.